This proposal is directed at understanding how the metabolism of trigonelline (N-methyl nicotinate) is regulated in relation to its function of promoting cell arrest in G2 in the cell cycle. Trigonelline is a component of the pyridine nucleotide pathway for the synthesis of NAD and NADP. Experiments are designed to understand how concentrations of nicotinic acid, nicotinamide, NAD, NADP, and other components of the pathway relate to trigonelline synthesis, degradation, and concentrations in relation to function. Trigonelline is present in cotyledons of ungerminated garden pea (Pisum sativum) seedlings and is transported to both roots and shoots after germination and promotes cell arrest in G2 of 40 and 10 percent of cells in roots and shoots, respectively, during normal cellular differentiation. This natural hormone (effective at 10 to the minus 7 M) directs cellular differentiation via cell cycle regulation. Experiments are planned to determine: a) concentration of four major components (trigonelline, nicotinic acid, nicotinamide and NAD) of the pathway in intact seedlings, b) structural groups of trigonelline required for function, c) rate of transport of trigonelline from cotyledons to roots, d) kinetics of trigonelline metabolism in root meristems at both effective and ineffective trigonelline concentrations in intact seedlings, e) range of plant species in which root meristems respond to trigonelline by showing prdominant cell arrest in G2. Quantities of trigonelline, nicotinic acid, nicotinamide, and NAD will be analyzed by high pressure liquid chromatography after separation and elution following thin-layer chromatography. Rates of conversion of trigonelline to other components of the pathway as well as rates of conversion of components to trigonelline will be determined with radioisotopes. Rates of conversion and quantities of components will be viewed in relation to cell cycle regulation. Preliminary results show that all legumes tested have large amounts of trigonelline in dry seeds while cereal grain crops (monocots) have little or none. Experiments are planned to determine if trigonelline is present in a wide variety of plants and if roots of such plants exhibit promotion of cell arrest in G2 if trigonelline is provided.